A New Surgical Technique for the Treatment of Congenital Concealed Penis Based on Anatomical Finding.

PURPOSE: Surgery is the first line treatment for congenital concealed penis but penile retraction is inevitable in some cases. We investigate the anatomical and histological characteristics of penile fasciae and describe a new technique for the correction of concealed penis. MATERIALS AND METHODS: The anatomical structures of penile fasciae were observed in 10 adult cadaveric penises. Penile tissue samples were stained with hematoxylin-eosin, Masson's trichrome and Weigert's resorcin-fuchsin, respectively. From January 2017 to May 2019, 78 patients with congenital concealed penis were treated with the new surgical technique. Median patient age was 14 years (range 8 to 18). RESULTS: Dartos fascia had sublayers. The superficial layer was a well vascularized tissue composed of nonpolar collagen fibers intermixed with nerves and vessels. The deep layer was composed of a transverse arrangement of collagen fibers and elastic fibers, and there were fewer venules and nerve fibers. Based on this finding we performed anatomical resection of the deep layer of dartos fascia to correct concealed penis. During the operation dartos fascia was separated into 2 layers and a complete circular resection of the deep layer was made at the base of the penis. Mean followup was 14 months. All patients and their parents were satisfied with the outcomes. None of the patients underwent postoperative penile retraction. CONCLUSIONS: The anatomical resection of the deep layer of dartos fascia for correcting concealed penis is technically easy, safe and effective. It provides a good cosmetic appearance and functional outcomes.

Early Aß-HBc virus-like particles immunization had better effects on preventing the deficit of learning and memory abilities and reducing cerebral Aß load in PDAPP mice.

For nearly two decades, immunization against the ß-amyloid peptide (Aß) has been investigated as a potential treatment for Alzheimer's disease (AD). Despite some disappointing results in clinic trials, greater significance has been attached by some researchers to exploring the immune effects on pathological and cognitive changes in AD or producing new vaccines of AD. In the previous study, we have made a virus-like particles (Aß-HBc VLPs) as Aß vaccine candidate. Aß-HBc VLPs could ameliorate the learning and memory abilities and reduce cerebral Aß deposit in the old PDAPP mice. In the present study, to observe the preventive effect and the proper time of immunization, 3, 6 and 9-month old PDAPP mice were immunized with Aß-HBc VLPs for 3 months. All mice generated high titer of anti-Aß antibody after Aß-HBc VLPs immunizations. When the mice were 15-month old, Morris Water Maze was used to test their learning and memory abilities. The escape latencies of Aß-HBc VLPs immunized mice were shorter than that of control mice. These immunized mice entered platform region frequently and spent more time on the platform region and quadrant. 3 m and 6 m Aß-HBc VLPs immunized groups performed better than the 9 m group. In immunohistochemistry tests, all the Aß-HBc VLPs immunized mice had less amyloid deposit in cortex and hippocampus. ELISA results showed that soluble Aß was reduced in the brain homogenates of the Aß-HBc VLPs immunized mice, and 3- and 6-month groups had less soluble Aß than the 9-month group. In conclusion, our study showed that Aß-HBc VLPs immunization could elicit a strong immune response in adult APP mice, and early immunization had better effects on preventing learning and memory deficits, lowering Aß burden in PDAPP mice.

Ablation of aberrant neurogenesis fails to attenuate cognitive deficit of chronically epileptic mice.

Pilocarpine-induced acute seizures strongly induce aberrant hippocampal neurogenesis, characterized by increased proliferation of neural progenitors and abnormal integrations of newly generated granule cells - hilar ectopic granule cells (EGCs), mossy fibre sprouting (MFS), and hilar basal dendrites (HBDs), which may disturb hippocampal neuronal circuits and thus contribute to cognitive impairment in temporal lobe epilepsy (TLE) patients and animal models. Previous studies via ablating hippocampal neurogenesis after acute seizures produced inconsistent results regarding the development of long-term cognitive impairment. Furthermore, a sufficient decrease of subsequent abnormal integrations in chronically epileptic hippocampus was not well-established in these studies. Therefore, the link between seizure-induced aberrant hippocampal neurogenesis and cognitive decline associated with epilepsy is still in need to be clarified. In this study, the mice were injected with methylazoxymethanol acetate (MAM) both before and after pilocarpine-induced status epilepticus (SE) to achieve an overall ablation of newborn cells contributing to the pathological recruitment. In addition, a protracted time point was chosen for behavioral testing considering it takes a fairly long time for newborn granule cells to adequately develop abnormal integrations, especially MFS. Although an overall reduction of abnormal integrations, including EGCs, MFS and HBDs was confirmed following the ablation regime, the performance of ablated and non-ablated mice in the Morris Water Maze (MWM) task did not differ. The current findings therefore provide novel evidences that ablation of neurogenesis with an overall decrease of abnormal integrations cannot attenuate subsequent cognitive impairment at least in the model used in this study.

Reduced abnormal integration of adult-generated granule cells does not attenuate spontaneous recurrent seizures in mice.

Epileptic seizures lead to aberrant hippocampal neurogenesis, including increased proliferation of neural progenitors and abnormal integrations of newly generated granule cells - hilar ectopic granule cells (EGCs), mossy fiber sprouting (MFS), and hilar basal dendrites (HBDs). Previous results from ablating hippocampal neurogenesis after acute seizures have been controversial with regards to the development of spontaneous recurrent seizures (SRSs). While ablation of hippocampal newborn cells was effective, a sufficient decrease of subsequent abnormal integrations in chronically epileptic hippocampus was not well-established in these studies. Evaluations of the role of aberrant neurogenesis in epileptogenesis were therefore inconclusive. In this study, we ablated the hippocampal neurogenesis by methylazoxymethanol acetate (MAM) treatment both before and after pilocarpine induced status epilepticus (SE). We found that an overall ablation of newborn granule cells and a protracted delay after the cell ablation are required to eliminate subsequent abnormal integrations, including EGCs, MSF and HBDs. However, there were no alterations in frequency, duration and severity of chronic seizures were demonstrated following this regime. The current findings provide novel evidences that an overall decrease of abnormal integrations via cell ablation cannot exert significant effects on the development of SRSs at least in the model used in this study.

Maternal separation exacerbates Alzheimer's disease-like behavioral and pathological changes in adult APPswe/PS1dE9 mice.

Alzheimer's disease (AD), the most common neurodegenerative disorder that gradually destroys memory and cognitive abilities in the elderly, makes a huge emotional and economic burden on the patients and their families. The presence of senile plaques and the loss of cholinergic neurons in the brain are two neuropathological hallmarks of AD. Maternal separation (MS) is an animal paradigm designed to make early life stress. Studies on wild type rodents showed that MS could induce AD-like cognitive deficit and pathological changes. However, the effects of MS on AD susceptible population or AD animal models are still unclear. In the present study, male APPswe/PS1dE9 transgenic mice were separated from dam and pups 3h per day from postnatal day 2 to day 21. After weaning, all animals were housed under normal conditions (4 mice per cage). At 9-month age, MWM tests were performed to evaluate the learning and memory abilities. Then the pathological changes in the brain were measured by histology staining. The results showed MS mice had more severe deficit of learning and memory. Compared to the control, there were more senile plaques in cortex and hippocampus, fewer cholinergic neurons in nucleus basalis of Meynert in MS mice. These results indicate that MS exacerbates Alzheimer's disease-like behavioral and pathological changes in APPswe/PS1dE9 mice.

Puerarin attenuates learning and memory impairments and inhibits oxidative stress in STZ-induced SAD mice.

Puerarin (PUE), an isoflavone purified from the root of Pueraria lobata (Chinese herb), has been reported to attenuate learning and memory impairments in the transgenic mouse model of Alzheimer's disease (AD). In the present study, we tested PUE in a sporadic AD (SAD) mouse model which was induced by the intracerebroventricular injection of streptozotocin (STZ). The mice were administrated PUE (25, 50, or 100mg/kg/d) for 28 days. Learning and memory abilities were assessed by the Morris water maze test. After behavioral test, the biochemical parameters of oxidative stress (glutathione peroxidase (GSH-Px), superoxide dismutases (SOD), and malondialdehyde (MDA)) were measured in the cerebral cortex and hippocampus. The SAD mice exhibited significantly decreased learning and memory ability, while PUE attenuated these impairments. The activities of GSH-Px and SOD were decreased while MDA was increased in the SAD animals. After PUE treatment, the activities of GSH-Px and SOD were elevated, and the level of MDA was decreased. The middle dose PUE was more effective than others. These results indicate that PUE attenuates learning and memory impairments and inhibits oxidative stress in STZ-induced SAD mice. PUE may be a promising therapeutic agent for SAD.

Vasoactive intestinal peptide administration after stroke in rats enhances neurogenesis and improves neurological function.

The aim of this study was to investigate the effects of vasoactive intestinal peptide (VIP) on neurogenesis and neurological function after cerebral ischemia. Rats were intracerebroventricular administered with VIP after a 2h middle cerebral artery occlusion (MCAO) and sacrificed at 7, 14 and 28 days after MCAO. Functional outcome was studied with the modified neurological severity score. The infarct volume was evaluated via histology. Neurogenesis, angiogenesis and the protein expression of vascular endothelial growth factor (VEGF) were measured by immunohistochemistry and Western blotting analysis, respectively. The treatment with VIP significantly reduced the neurological severity score and the infarc volume, and increased the numbers of bromodeoxyuridine (BrdU) immunoreactive cells and doublecortin immunoreactive area in the subventricular zone (SVZ) at 7, 14 and 28 days after ischemia. The cerebral protein levels of VEGF and VEGF expression in the SVZ were also enhanced in VIP-treated rats at 7 days after stroke. VIP treatment obviously increased the number of BrdU positive endothelial cells in the SVZ and density of cerebral microvessels in the ischemic boundary at 28 days after ischemia. Our study suggests that in the ischemic rat brain VIP reduces brain damage and promotes neurogenesis by increasing VEGF. VIP-enhanced neurogenesis is associated with angiogenesis. These changes may contribute to improvement in functional outcome.

Mitogen-activated protein kinase signaling pathways promote low-density lipoprotein receptor-related protein 1-mediated internalization of beta-amyloid protein in primary cortical neurons.

Mounting evidence suggests that the pathological hallmarks of Alzheimer's disease (AD) are caused by the intraneuronal accumulation of beta-amyloid protein (Aß). Reuptake of extracellular Aß is believed to contribute significantly to the intraneuronal Aß pool in the early stages of AD. Published reports have claimed that the low-density lipoprotein receptor-related protein 1 (LRP1) mediates Aß1-42 uptake and lysosomal trafficking in GT1-7 neuronal cells and mouse embryonic fibroblast non-neuronal cells. However, there is no direct evidence supporting the role of LRP1 in Aß internalization in primary neurons. Our recent study indicated that p38 MAPK and ERK1/2 signaling pathways are involved in regulating α7 nicotinic acetylcholine receptor (α7nAChR)-mediated Aß1-42 uptake in SH-SY5Y cells. This study was designed to explore the regulation of MAPK signaling pathways on LRP1-mediated Aß internalization in neurons. We found that extracellular Aß1-42 oligomers could be internalized into endosomes/lysosomes and mitochondria in cortical neurons. Aß1-42 and LRP1 were also found co-localized in neurons during Aß1-42 internalization, and they could form Aß1-42-LRP1 complex. Knockdown of LRP1 expression significantly decreased neuronal Aß1-42 internalization. Finally, we identified that p38 MAPK and ERK1/2 signaling pathways regulated the internalization of Aß1-42 via LRP1. Therefore, these results demonstrated that LRP1, p38 MAPK and ERK1/2 mediated the internalization of Aß1-42 in neurons and provided evidence that blockade of LRP1 or inhibitions of MAPK signaling pathways might be a potential approach to lowering brain Aß levels and served a potential therapeutic target for AD.

[Optimization of trypsin digestion intensity to obtain high-purity in vitro cultured astrocytes].

The aim of the present study was to observe the effect of trypsin digestion on the purity of in vitro cultured astrocytes and optimize the culture methods. The cerebral cortical tissue from newborn Sprague Dawley (SD) rats was isolated and digested with 0.25% trypsin for 20, 30, or 40 min. The obtained single cell suspension was then cultured. Once reaching confluence, the cells were shaken at a constant temperature. Then, each of 20 and 30 min groups was subdivided into two groups, the control group with normal digestion and two-time-digestion group, and the cells were passaged and purified. Through inverted phase contrast microscope and MTT assay, cell growth and proliferation were observed, respectively. Immunofluorescence for glial fibrillary acidic protein (GFAP) was used to observe the morphology of astrocytes and to assess their purity in different stages. Flow cytometric analysis was used to detect the apoptotic rates of purified astrocytes. The results showed that, the cells being digested for 20 min usually reached confluence at 9 d after seeding. When the digestion time was extended to 30 min, the cells grew faster and reached confluence at 7 d after seeding, meanwhile the morphology of astrocytes was normal, GFAP positive rate (70.2 ± 4.0)% being much higher than that of the 20 min group (P < 0.05). Compared with 20 min group, 40 min group showed higher GFAP positive rate, whereas the cell proliferation was slower, and cell injury was more obvious. After shaking at constant temperature, two times of trypsin digestion could decrease the number of contaminated cells after passage. The GFAP positive rates of two-time-digestion groups in passage 1 (P1) were higher than those of corresponding control groups, and the GFAP positive rate of 30 min + two-time-digestion group in P1 reached (98.1 ± 1.7)%, which was equivalent to that of the 20 min + control group in P3. However, the apoptotic rate showed no significant difference between these two groups. Based on above mentioned results, we conclude that 30 min + two-time of trypsin digestion effectively improves the purity of astrocytes and shortens the time of primary culture and purification, suggesting that it is a rapid and effective method to obtain astrocytes with high purity in vitro.

Aß-HBc virus-like particles immunization without additional adjuvant ameliorates the learning and memory and reduces Aß deposit in PDAPP mice.

ß-Amyloid peptide (Aß) immunization is regarded as a promising therapy to Alzheimer's disease. The full length Aß as antigen might induce meningoencephalitis adverse effect since the middle and C-terminal fragments of Aß contain T cell epitopes. While N-terminal fragment of Aß, containing B cell epitope, has weak or no immunogenicity. To improve the immunogenicity, in the previous study, we used HBv core antigen as carrier to make fusion protein containing 2 Aß1-15. The fusion protein could form virus-like particles (VLPs) and had strong immunogenicity. The antisera prevented Aß fiber formation and protected the PC12 cells against toxicity of Aß. In the present study, we immunized 12-month old AD transgenic mice, PDAPP mice, to observe the therapeutic effect of immunization on behaviour and pathology. During immunization, the titer of anti-Aß antibody reached to nearly 1:10(6) after 4th inoculation, and then maintained that level to the end of the experiment. After 6-month immunization, the behavioral changes of mice were tested by Morris Water Maze (MWM). The escape latency of immunized mice was shorter than control, and these mice entered platform quadrant more times. Immunohistochemistry results showed that Aß-HBc VLPs immunized mice had less amyloid deposit with less microglia in cortex and hippocampus. In conclusion, Aß-HBc VLPs ameliorated the learning and memory and reduced cerebral Aß deposit in PDAPP mice.

Taurine attenuates amyloid ß 1-42-induced mitochondrial dysfunction by activating of SIRT1 in SK-N-SH cells.

Amyloid ß (Aß) plays a critical role in the pathogenesis of Alzheimer disease (AD). Studies indicate that Aß causes reactive oxygen species (ROS) generation, mitochondrial dysfunction and neurons loss in vivo and in vitro. Taurine, a naturally occurring ß-amino acid in the brain, has been demonstrated to have neuroprotective properties. In the present study, the effects of taurine on cell viability and mitochondrial function in Aß1-42-treated SK-N-SH cells were investigated. Pretreatment of taurine significantly attenuated Aß1-42-induced neuronal death. Similarly, taurine suppressed the mPTP opening and reversed mitochondrial function in the presence of Aß1-42. Additionally, taurine attenuated the intracellular Ca(2+) and ROS generation induced by Aß1-42. Moreover, the expression of Sirtuin 1 (SIRT1) was obviously recovered by taurine in Aß1-42-treated SK-N-SH cells. Our results suggest that taurine prevents Aß1-42-induced mitochondrial dysfunction by activation of SIRT1. This study implies that taurine is a prospective additive for AD patients.

The effects of valsartan on cognitive deficits induced by aluminum trichloride and d-galactose in mice.

OBJECTIVES: Valsartan has been reported to reduce brain beta-amyloid protein levels and improve spatial learning in the Tg2576 transgenic mouse model of Alzheimer's disease (AD). However, the exact mechanism of neuroprotective effects of valsartan has not been properly studied especially in cholinergic function and oxidative damage, the essential factors that undergo impairment in AD. Therefore, the present study examined the effects of valsartan on memory impairment, cholinergic dysfunction, and oxidative stress in aluminum trichloride (AlCl3) and d-galactose (d-gal)-induced experimental sporadic dementia of Alzheimer's type. METHODS: Valsartan was administered intragastrically (i.g.) (20 mg/kg/day) for 60 days after mice were given AlCl3 (10 mg/kg/day) and d-gal (150 mg/kg/day) intraperitoneally (i.p.) once daily for 90 days. Then, memory function was evaluated by Morris water maze test. Acetylcholinesterase (AChE), superoxide dismutases (SOD) and glutathione peroxidase (GSH-Px) activities and malondialdehyde (MDA) level in cortex and hippocampus were also assessed with biochemical technique. RESULTS: Chronic administration of valsartan not only improved learning and memory but also restored the elevation of AChE activity induced by AlCl3 and d-gal in cortex and hippocampus. In addition, valsartan significantly restored SOD and GSH-Px activities and reduced MDA level in cortex and hippocampus indicating attenuation of oxidative stress. DISCUSSION: Our results indicate that valsartan prevents AlCl3- and d-gal-induced cognitive decline partly to restore cholinergic function and attenuate oxidative damage. These findings further support the potential of valsartan to be used in AD treatment.

The effects of perindopril on cognitive impairment induced by d-galactose and aluminum trichloride via inhibition of acetylcholinesterase activity and oxidative stress.

Preclinical and clinical studies indicate involvement of renin angiotensin system (RAS) in memory functions. However, exact role of RAS in cognition is still ambiguous. The present study investigated the effects of perindopril on dementia of Alzheimer's type induced by d-galactose (d-gal) and aluminum trichloride (AlCl3). Perindopril, an angiotensin converting enzyme inhibitor, was administered intragastrically (0.5mg/kg/day) for 60days after mice were given d-gal (150mg/kg/day) and AlCl3 (10mg/kg/day) intraperitoneally for 90days. Then, memory function was evaluated by Morris water maze test. The biochemical studies were conducted in cerebral cortex and hippocampus of mouse brain after the behavioral studies. d-Gal and AlCl3 caused significant memory impairment along with significant elevation of acetylcholinesterase (AChE) activity in cerebral cortex and hippocampus. Further, a significant reduction of superoxide dismutases (SOD) and glutathione peroxidase (GSH-Px) activities, and elevation of malondialdehyde (MDA) level in cerebral cortex and hippocampus were observed. Perindopril not only improved cognitive impairment but also restored the elevation of AChE activity induced by d-gal and AlCl3. In addition, perindopril significantly increased SOD and GSH-Px activities, reduced MDA level in cerebral cortex and hippocampus. Taken together, the above findings indicate that perindopril improves learning and memorizing probably by restoring cholinergic function and attenuating oxidative damage.

Anti-Aß antibodies induced by Aß-HBc virus-like particles prevent Aß aggregation and protect PC12 cells against toxicity of Aß1-40.

ß-Amyloid peptide (Aß) immunization is regarded as the most promising therapy to Alzheimer' s disease. The full length Aß as antigen might induce meningoencepholontis adverse effect since the middle and C-terminal fragments of Aß contain T cell epitopes. While N-terminal fragment of Aß, containing B cell epitope, has weak or no immunogenicity. To improve the immunogenicity, we used HBV core antigen as carrier to make fusion protein containing 2 Aß1-15. The fusion protein was expressed in Escherichia coli harboring the recombinant plasmid pET/c-2Aß15-c. Transmission electron microscope (TEM) showed that fusion protein could form virus-like particles (VLPs). After 7-weeks immunization with Aß-HBc VLPs through subcutaneous injection, the titer of anti-Aß antibody in sera of BALB/c mice reached up to 10(5), higher than Aß peptide immunization. Aß-HBc VLPs immunization did not elicit Aß-specific T cell proliferation. The main isotypes of antibody in mice immunized with Aß-HBc VLPs were IgG1 and IgG2b, while isotype in mice immunized with Aß1-42 was IgG2a. When the antisera from mice immunized with Aß-HBc VLPs were co-incubated for 1 week at 37°C with Aß, fibers of aggregated Aß was reduced or diminished. The antibodies also prevented PC12 cells from injury by toxicity of Aß. In conclusion, recombinant c-2Aß15-c gene can be expressed in E. coli. The expressed protein could form VLPs and has strong immunogenicity. The antisera prevented Aß fiber formation and protected the PC12 cells against toxicity of Aß. This study lays the foundation for the experimental study of AD gene engineering vaccine.

Vasoactive intestinal peptide increases VEGF expression to promote proliferation of brain vascular endothelial cells via the cAMP/PKA pathway after ischemic insult in vitro.

Vasoactive intestinal peptide (VIP) enhances angiogenesis in rats with focal cerebral ischemia. In the present study, we investigated the molecular mechanism of the proangiogenic action of VIP using an in vitro ischemic model, in which rat brain microvascular endothelial cells (RBMECs) are subjected to oxygen and glucose deprivation (OGD). Western blotting and immunocytochemistry were carried out to examine the expression of VIP receptors and vascular endothelial growth factor (VEGF) in cultured RBMECs. The cell proliferation was assessed by the MTT assay. Cyclic adenosine monophosphate (cAMP) and VEGF levels were measured by using the enzyme-linked immunosorbent assay. The cultured RBMECs expressed VPAC1, VPAC2 and PAC1 receptors. Treatment with VIP significantly promoted the proliferation of RBMECs and increased OGD-induced expression of VEGF, and this effect was antagonized by the VPAC receptor antagonist VIP6-28 and VEGF antibody. VIP significantly increased contents of cAMP in RBMECs and VEGF in the culture medium. The VIP-induced VEGF production was blocked by H89, a protein kinase A (PKA) inhibitor. These data suggest that treatment with VIP promotes VEGF-mediated endothelial cell proliferation after ischemic insult in vitro, and this effect appears to be initiated by the VPAC receptors leading to activation of the cAMP/PKA pathway.

[Prokaryotic expression and immunogenicity of the chimeric HBcAg containing Aß(1-15)].

OBJECTIVE: To construct a recombinant prokaryoticexpression plasmid pET/ c-Aß(15)-c, and evaluate the immunogenicity of its encoded fusion protein as expressed in E.coli. METHODS: The gene fragment HBc88-144 was amplified by PCR and subcloned to pUC19. The synthetic, double-strand Aß(1-15) gene was inserted downstream of HBc1-71 in pGEMEX/c1-71. After restriction enzyme digestion, c1-71- Aß(15) was spliced to HBc88-144, yielding the recombinant gene c-Aß(15)-c; that gene was subcloned into pET-28a(+). The fusion protein (CA15C) expressed in the transformed E.coli BL21 was induced with isopropyl ß-D-1-thiogalactopyranoside (IPTG) and analyzed by SDS-PAGE. The virus-like particle (VLP) formed by fusion protein CA15C was observed with transmission electric microscope (TEM). Four Kunming (KM) mice were given intraperitoneal injections of CA15C, and the anti-Aß antibody elicited was detected by indirect ELISA. RESULTS: The sequence of the recombinant gene was confirmed by restriction enzyme digestion and DNA sequencing. After IPTG induction, the fusion protein was expressed, mainly in the sediment from the bacterial lysate. The expression level was 40% of total protein in the sediment. The CA15C could form VLP. After 5 rounds of immunization, the titer of anti-Aß antibody in the sera of KM mice reached 1:10000, while the anti-HBc antibody was undetectable. CONCLUSION: Recombinant c-Aß(15)-c gene can be expressed in E.coli. The expressed protein can form VLPs and has a strong immunogenicity.

Association between the apolipoprotein E4 and postoperative cognitive dysfunction in elderly patients undergoing intravenous anesthesia and inhalation anesthesia.

BACKGROUND: Intravenous and inhalation anesthesia are commonly used in the clinical setting. Recovery of cognitive function in elderly patients after surgery has received increased attention. In this study, the authors compared recovery of cognitive function in patients after different anesthesia techniques, and investigated which technique is safer. The authors also explored association between apolipoprotein E4 and postoperative cognitive dysfunction in patients undergoing general anesthesia. METHODS: A total of 2,000 patients were equally and randomly divided into intravenous and inhalation anesthesia groups. Total intravenous and inhalation anesthesia were used. Within 10 days after surgery, cognitive function was assessed daily using the Mini-Mental State Examination (MMSE). Restriction fragment length polymorphism of apolipoprotein E gene was analyzed. The primary outcome was MMSE score, frequency distribution of apolipoprotein E alleles and genotypes. P < 0.01 was used as statistically significant. RESULTS: MMSE score in inhalation preoperative baseline group significantly decreased at day 3 after surgery compared with the preoperational and intravenous anesthesia group. The proportion of patients scoring less than 25 points was significantly greater in the inhalation anesthesia group than in the intravenous anesthesia group at 3 days after surgery. In the inhalation anesthesia group, the decrease in MMSE score was closely related with apolipoprotein E ε4 allele. In the intravenous anesthesia group, the decrease in MMSE score was not correlated with apolipoprotein E ε4 allele. CONCLUSIONS: There was a strong association between the apolipoprotein E ε4 and postoperative cognitive dysfunction in elderly patients undergoing inhalation anesthetics.

[Prokaryotic expression and immunogenicity analysis of the chimeric HBcAg containing APP beta cleavage site peptide and Aß(1-15);].

AIM: To construct the recombinant prokaryotic expression plasmid pET/c-ABCSP-Aß(15-c);, and evaluate the immunogenicity of the fusion protein expressed in E.coli. METHODS: The gene fragment HBc88-144 was amplified by PCR and subcloned to pUC19. The APP beta cleavage site peptide(ABCSP) and Aß(1-15); gene(ABCSP-Aß(15);) was amplified by PCR and inserted downstream of HBc1-71 in pGEMEX/c1-71. After restriction enzyme digestion, c1-17-ABCSP-Aß(15); were connected with HBc88-144, yielding the recombinant gene c-ABCSP-Aß(15-c);. c-ABCSP-Aß(15-c); gene was subcloned into pET-28a(+).The fusion protein expressed in transformed E.coli BL21 was induced with IPTG and analyzed by SDS-PAGE. The virus-like particles (VLP) formed by fusion protein was observed with Transmission Electron Microscope (TEM). 4 Kunming (KM) mice received intraperitoneal injection (i.p) of fusion protein VLP. The antibody was detected by indirect ELISA. RESULTS: The recombinant gene was confirmed by restriction enzyme digestion and DNA sequencing. After IPTG induction, fusion protein was expressed and mainly existed in the sediment of the bacterial lysate. The expression level was 40% of all the proteins in the sediment. The fusion protein could form VLP. After 5 times of immunization, the titer of anti-ABCSP and anti-Aßantibody in sera of KM mice reached up to 1:5 000 and 1:10 000 respectively, while the anti-HBc antibody was undetectable. CONCLUSION: Recombinant c-ABCSP-Aß(15-c); gene can be expressed in E.coli. The expressed protein could form VLP and has a strong immunogenicity. This study lays the foundation for the study of AD genetic engineering vaccine.

[Immunization with recombinant HBcAg and ß-amyloid peptide fusion protein promotes clearance of intrahippocampally injected ß-amyloid peptide in rats].

OBJECTIVE: To study the effects of immunization with the fusion protein CAC (a product of prokaryotic expression of recombinant HBcAg and ß-amyloid peptide fusion gene) against the toxicity induced by intrahippocampal injection of aggregated ß-amyloid peptide (Aß) in rats. METHODS: SD rats were immunized intraperitoneally with the fusion protein CAC, and the titer of anti-Aß antibody was evaluated by ELISA. When the titers of the anti-Aß antibody reached 1:3 000, aggregated Aß was injected into the CA1 region of the rat hippocampus. Two weeks after Aß injection, the rats underwent morris water maze test before sacrificed to prepare the brain slices with Congo red and haematoxylin staining. RESULTS: The titer of anti-Aß antibody reached 1:3 000 after 5 immunizations with the fusion protein. After Aß injection, the saline-immunized rats showed a reduced cognitive behavior in the Morris water maze test compared to the CAC-immunized rats. In the saline-immunized rats, the neurons around the site of Aß injection exhibited obvious cell damages with Aß deposits and glial infiltration, whereas in CAC-immunized rats, Aß deposits were significantly reduced or even absent. CONCLUSION: Immunization with the fusion protein CAC can inhibit the toxicity induced by intrahippocampal aggregated Aß injection.